Vitrification of murine mature metaphase II oocytes perturbs DNA methylation reprogramming during preimplantation embryo development

Cao, Zubing; Zhang, Meiling; Xu, Tengteng; Chen, Zhen; Xu, Tong; Zhang, Dandan; Wang, Yiqing; Zhang, Ling; Gao, Di; Luo, Lei; Khan, Ibrar Muhammad

doi:10.1016/j.cryobiol.2019.01.012

Cited by 11 publications

(8 citation statements)

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“…As expected, 29,39,40 DNA methylation and hydroxymethylation levels gradually decreased during the in vivo development of embryos (Figure 6A,B). However, this dynamic was not found in the embryos obtained after in vitro culture of zygotes or after ICSI.…”

Section: Discussionsupporting

confidence: 82%

Dynamics of epigenetic modifications in ICSI embryos from in vitro‐produced spermatozoa

et al. 2020

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Background In prepubertal boys with cancer, fertility preservation relies on testicular tissue freezing before treatment. In vitro maturation of frozen/thawed tissues could be one of the procedures envisaged to restore the fertility of cured patients. It is necessary to ascertain in the mouse model that in vitro‐generated spermatozoa are able to ensure embryo development, without altering the epigenetic processes occurring during the pre‐implantation period. Objectives The aims of the present study were to investigate the fertilizing ability of in vitro‐produced spermatozoa and explore several epigenetic marks at different stages of embryo development. Materials and methods Fresh or controlled slow‐frozen (CSF)/thawed testicular tissues from 6 to 7 days post‐partum (dpp) mice were cultured for 30 days. Intracytoplasmic sperm injection (ICSI) experiments were performed using in vitro‐produced spermatozoa. Testicular spermatozoa from 36 to 37 dpp mice were used as in vivo controls. DNA methylation/hydroxymethylation and histone post‐translational modifications (H3K4me3, H3K27me3 and H3K9ac) were analysed by immunofluorescence from the zygote to the blastocyst stages. Results The spermatozoa generated in cultures of fresh or CSF testicular tissues were able to initiate embryonic development. The freezing of prepubertal testicular tissues limits the production of spermatozoa in vitro and the fertilization rate after ICSI. Similar levels of H3K4me3, H3K27me3 and H3K9ac were found in ICSI embryos derived from in vitro‐ and in vivo‐produced spermatozoa. DNA methylation levels were increased in 4‐cell embryos and morula obtained by ICSI with in vitro‐produced spermatozoa. Discussion and conclusion Our study shows for the first time that the use of in vitro‐produced spermatozoa alters DNA methylation/demethylation dynamics but has little impact on H3K4me3, H3K27me3 and H3K9ac levels in mouse early embryos. Further work will have to be performed to determine whether the use of these gametes is not deleterious for embryo development before considering a human application.

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Section: Discussionsupporting

confidence: 82%

Dynamics of epigenetic modifications in ICSI embryos from in vitro‐produced spermatozoa

et al. 2020

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“…In recent years, the quest to improve oocyte cryopreservation protocols has remained central in the field of reproductive medicine, which seeks to provide optimal [142,143], pig [144], cattle [112], and sheep models [141], as well as reduced maturation rates in vitrified immature human oocytes [145]. Additional studies have reported ultrastructural, biochemical, and molecular changes as a result of oocyte vitrification [146].…”

Section: Discussionmentioning

confidence: 99%

“…To date, although vitrification protocols result in relative high survival rates, they do not necessarily signify developmental competence. Previous reports highlighted a decreased cleavage and blastocyst rates after oocyte vitrification in mouse [ 142 , 143 ], pig [ 144 ], cattle [ 112 ], and sheep models [ 141 ], as well as reduced maturation rates in vitrified immature human oocytes [ 145 ]. Additional studies have reported ultrastructural, biochemical, and molecular changes as a result of oocyte vitrification [ 146 ].…”

Section: Discussionmentioning

confidence: 99%

What impact does oocyte vitrification have on epigenetics and gene expression?

Barberet¹,

Barry²,

Choux³

et al. 2020

Clin Epigenet

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Children conceived by assisted reproductive technologies (ART) have a moderate risk for a number of adverse events and conditions. The question whether this additional risk is associated with specific procedures used in ART or whether it is related to the intrinsic biological factors associated with infertility remains unresolved. One of the main hypotheses is that laboratory procedures could have an effect on the epigenome of gametes and embryos. This suspicion is linked to the fact that ART procedures occur precisely during the period when there are major changes in the organization of the epigenome. Oocyte freezing protocols are generally considered safe; however, some evidence suggests that vitrification may be associated with modifications of the epigenetic marks. In this manuscript, after describing the main changes that occur during epigenetic reprogramming, we will provide current information regarding the impact of oocyte vitrification on epigenetic regulation and the consequences on gene expression, both in animals and humans. Overall, the literature suggests that epigenetic and transcriptomic profiles are sensitive to the stress induced by oocyte vitrification, and it also underlines the need to improve our knowledge in this field.

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“…Medically assisted reproduction procedures may subject gametes and early embryos to environmental stress which may impact the development of the foetus, for example, by disruptions in the expression of imprinted genes ( Sakian et al , 2015 ). Gamete and embryo cryopreservation methods can also affect DNA methylation ( Van Heertum and Weinerman, 2018 ; Cao et al , 2019 ).…”

Section: Introductionmentioning

confidence: 99%

Association of medically assisted reproduction with offspring cord blood DNA methylation across cohorts

et al. 2021

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STUDY QUESTION Is cord blood DNA methylation associated with having been conceived by medically assisted reproduction? SUMMARY ANSWER This study does not provide strong evidence of an association of conception by medically assisted reproduction with variation in infant blood cell DNA methylation. WHAT IS KNOWN ALREADY Medically assisted reproduction consists of procedures used to help infertile/subfertile couples conceive, including ART. Due to its importance in gene regulation during early development programming, DNA methylation and its perturbations associated with medically assisted reproduction could reveal new insights into the biological effects of assisted reproductive technologies and potential adverse offspring outcomes. STUDY DESIGN, SIZE, DURATION We investigated the association of DNA methylation and medically assisted reproduction using a case–control study design (N = 205 medically assisted reproduction cases and N = 2439 naturally conceived controls in discovery cohorts; N = 149 ART cases and N = 58 non-ART controls in replication cohort). PARTICIPANTS/MATERIALS, SETTINGS, METHODS We assessed the association between medically assisted reproduction and DNA methylation at birth in cord blood (205 medically assisted conceptions and 2439 naturally conceived controls) at >450 000 CpG sites across the genome in two sub-samples of the UK Avon Longitudinal Study of Parents and Children (ALSPAC) and two sub-samples of the Norwegian Mother, Father and Child Cohort Study (MoBa) by meta-analysis. We explored replication of findings in the Australian Clinical review of the Health of adults conceived following Assisted Reproductive Technologies (CHART) study (N = 149 ART conceptions and N = 58 controls). MAIN RESULTS AND THE ROLE OF CHANCE The ALSPAC and MoBa meta-analysis revealed evidence of association between conception by medically assisted reproduction and DNA methylation (false-discovery-rate-corrected P-value < 0.05) at five CpG sites which are annotated to two genes (percentage difference in methylation per CpG, cg24051276: Beta = 0.23 (95% CI 0.15,0.31); cg00012522: Beta = 0.47 (95% CI 0.31, 0.63); cg17855264: Beta = 0.31 (95% CI 0.20, 0.43); cg17132421: Beta = 0.30 (95% CI 0.18, 0.42); cg18529845: Beta = 0.41 (95% CI 0.25, 0.57)). Methylation at three of these sites has been previously linked to cancer, aging, HIV infection and neurological diseases. None of these associations replicated in the CHART cohort. There was evidence of a functional role of medically assisted reproduction-induced hypermethylation at CpG sites located within regulatory regions as shown by putative transcription factor binding and chromatin remodelling. LIMITATIONS, REASONS FOR CAUTIONS While insufficient power is likely, heterogeneity in types of medically assisted reproduction procedures and between populations may also contribute. Larger studies might identify replicable variation in DNA methylation at birth due to medically assisted reproduction. WIDER IMPLICATIONS OF THE FINDINGS Newborns conceived with medically assisted procedures present with divergent DNA methylation in cord blood white cells. If these associations are true and causal, they might have long-term consequences for offspring health. STUDY FUNDING/COMPETING INTERESTS(S) This study has been supported by the US National Institute of Health (R01 DK10324), the European Research Council under the European Union’s Seventh Framework Programme (FP7/2007-2013)/ERC Grant agreement no. 669545, European Union’s Horizon 2020 research and innovation programme under Grant agreement no. 733206 (LifeCycle) and the NIHR Biomedical Centre at the University Hospitals Bristol NHS Foundation Trust and the University of Bristol. The UK Medical Research Council and Wellcome (Grant ref: 102215/2/13/2) and the University of Bristol provide core support for ALSPAC. Methylation data in the ALSPAC cohort were generated as part of the UK BBSRC funded (BB/I025751/1 and BB/I025263/1) Accessible Resource for Integrated Epigenomic Studies (ARIES, http://www.ariesepigenomics.org.uk). D.C., J.J., C.L.R. D.A.L and H.R.E. work in a Unit that is supported by the University of Bristol and the UK Medical Research Council (Grant nos. MC_UU_00011/1, MC_UU_00011/5 and MC_UU_00011/6). B.N. is supported by an NHMRC (Australia) Investigator Grant (1173314). ALSPAC GWAS data were generated by Sample Logistics and Genotyping Facilities at Wellcome Sanger Institute and LabCorp (Laboratory Corporation of America) using support from 23andMe. The Norwegian Mother, Father and Child Cohort Study is supported by the Norwegian Ministry of Health and Care Services and the Ministry of Education and Research, NIH/NIEHS (Contract no. N01-ES-75558), NIH/NINDS (Grant nos. (i) UO1 NS 047537-01 and (ii) UO1 NS 047537-06A1). For this work, MoBa 1 and 2 were supported by the Intramural Research Program of the NIH, National Institute of Environmental Health Sciences (Z01-ES-49019) and the Norwegian Research Council/BIOBANK (Grant no. 221097). This work was partly supported by the Research Council of Norway through its Centres of Excellence funding scheme, Project no. 262700. D.A.L. has received support from national and international government and charity funders, as well as from Roche Diagnostics and Medtronic for research unrelated to this study. The other authors declare no conflicts of interest. TRIAL REGISTRATION NUMBER N/A.

show abstract

Vitrification of murine mature metaphase II oocytes perturbs DNA methylation reprogramming during preimplantation embryo development

Cited by 11 publications

References 30 publications

Dynamics of epigenetic modifications in ICSI embryos from in vitro‐produced spermatozoa

Dynamics of epigenetic modifications in ICSI embryos from in vitro‐produced spermatozoa

What impact does oocyte vitrification have on epigenetics and gene expression?

Association of medically assisted reproduction with offspring cord blood DNA methylation across cohorts

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